Phenotypic and Genotypic Characterization of Neisseria gonorrhoeae Isolates from New Zealand with Reduced Susceptibility to Ceftriaxone.

Aim: To characterize mutations in penA, mtrR, ponA, and porBIB, considered target genes for antimicrobial resistance, in Neisseria gonorrhoeae isolates with elevated minimum inhibitory concentrations (MICs) of ceftriaxone cultured from patients in New Zealand. Results: Out of 28 isolates supplied by the Institute of Environmental Science and Research Limited (ESR), Porirua, New Zealand, 14 were found to show reduced susceptibility to ceftriaxone (MIC of 0.06 mg/L) according to criteria used by the ESR and the Australian Gonococcal Surveillance Programme (AGSP) when tested in our laboratory. Rates of resistance to ciprofloxacin, azithromycin, penicillin, and tetracycline were 100% (28/28), 7% (2/28), 36% (10/28), and 25% (7/28), respectively. Ten different penA (Penicillin binding protein 2 [PBP2]) sequences were observed. The most common mosaic penA M-1 resembled mosaic penA XXXIV, which has been associated with ceftriaxone treatment failures in other countries. Four semimosaic PBP2 sequences were observed and may be novel PBP sequences, while four out of five nonmosaic PBP2 sequences were similar to PBP2 sequences reported in Australia. Twenty-one isolates harbored mutations in all 4 genes (penA, mtrR, porBIB, and ponA), and 13 of these exhibited reduced susceptibility to ceftriaxone. Conclusion: Mutations in penA, mtrR, porBIB, and ponA observed in this study may have contributed to reduced susceptibility to ceftriaxone among New Zealand gonococcal isolates. Over half (16/22) of mosaic penA sequences from the gonococcal isolates resembled penA XXXIV.

Emm type distribution of group A streptococcus isolates from the throat swabs of children living in areas with a high (Northland and Gisborne) or low (Palmerston North) incidence of acute rheumatic fever.

AIM: To assess the circulating emm types of pharyngeal isolates of group A streptococcus (GAS) among school children living in Northland, the Gisborne region and Palmerston North, New Zealand. METHODS: GAS were isolated from throat swabs sent to laboratories in Northland (197 in 2013) and Gisborne (115 in 2014-15) and from children enrolled in the Palmerston North Solar Ventilation Project (70 in 2013-14). The incidences of acute rheumatic fever (ARF) cases in the three regions in 2014 were 9, 19.1 and 0 cases per 100,000 for Northland, the Gisborne region and Palmerston North respectively. DNA sequencing of the N-terminal portion of the emm gene was performed at the Institute of Environmental Science and Research Limited (ESR) laboratory (Porirua, New Zealand). RESULTS: A total of 36 emm types were found among pharyngeal GAS isolates from Northland children with emm1 predominating (24%), 28 emm types from the Gisborne region with emm12 predominating (25%) and 20 emm types from Palmerston North, again with emm12 predominating (36%). Of these GAS isolates, 38% were emm pattern A-C, usually associated with throat infections, 23% were pattern D, usually associated with skin infections, and 39% pattern E or generalists. The most common of the 13 emm clusters detected were A-C4 (emm12; 18% isolates), A-C3 (emm1, emm227, emm238; 17% isolates), D4 (9 emm types; 16% isolates), E4 and E3 (8 emm types each; 15% and 10% isolates respectively). A total of 301 of the 376 (80%) isolates were serotypes previously associated with ARF in New Zealand. CONCLUSION: The only significant differences in distribution between the regions with high (Northland and Gisborne area) and low (Palmerston North) incidences of ARF were the presence of emm3 and absence of emm41 among GAS isolates from Palmerston North school children.

Direct detection of markers associated with Neisseria gonorrhoeae antimicrobial resistance in New Zealand using residual DNA from the Cobas 4800 CT/NG NAAT assay.

OBJECTIVES: To use nucleic acid amplification techniques (NAAT) for detection of markers associated with gonococcal antimicrobial resistance (AMR) in non-cultured clinical samples to enhance surveillance of Neisseria gonorrhoeae AMR in New Zealand. METHODS: A total of 198 clinical samples from patients living in two cities, Wellington and Auckland and the more rural region of Gisborne, New Zealand, which were positive for N. gonorrhoeae by the Cobas 4800 were tested for three markers that predict reduced susceptibility or resistance to three antibiotics. Residual DNA extracts from the Cobas 4800 NG/CT test were tested for a single-nucleotide polymorphism in the gyrA gene at codon 91 associated with quinolone resistance; a sequence on the plasmid in penicillinase-producing N. gonorrhoeae (PPNG) which confers resistance to penicillin and the mosaic penA sequence associated with reduced susceptibility to extended-spectrum cephalosporins in N. gonorrhoeae. RESULTS: A total of 186/198 (94%) of the samples provided a valid result on gyrA genotyping, confirming the utility of N. gonorrhoeae DNA extracted by the Roche Cobas 4800 CT/NG test for subsequent detection of AMR markers. The NAAT results for Wellington, Auckland and Gisborne, respectively, showed that 77%, 33% and 32% of samples had the marker associated with quinolone resistance, while 4%, 15% and 0% were positive for the PPNG plasmid marker, and 9%, 5% and 0% samples were positive for mosaic penA sequence. CONCLUSIONS: The use of residual clinical DNA samples from the Cobas 4800 CT/NG test proved an efficient and effective method for performing AMR genotyping. These data also show for the first time the presence of gonococci with a mosaic penA sequence in New Zealand. Overall, the results further highlight the potential of molecular methods to aid N. gonorrhoeae AMR surveillance, particularly for regions where gonococcal culture is no longer performed.

Arcobacter species in diarrhoeal faeces from humans in New Zealand.

AIM: To determine the prevalence, genetic diversity and antimicrobial susceptibility of Arcobacter spp in faecal samples from humans with diarrhoea in New Zealand. METHODS: An enrichment method was used to isolate Arcobacter spp from diarrhoeal human faeces submitted to a community laboratory in Hawke's Bay. The identity of isolates was confirmed by PCR and their diversity was determined by pulsed field gel electrophoresis (PFGE). Antibiotic susceptibility was established with E test strips. RESULTS: Arcobacter spp were isolated from 12 of 1380 diarrhoeal faecal samples examined (0.9%), including 7 A. butzleri and 5 A. cryaerophilus. Additional enteric pathogens were detected in four of these diarrhoeal faecal samples. All the Arcobacter isolates were genetically distinct and susceptible to ciprofloxacin. Most were also susceptible to erythromycin (92%) but fewer to tetracycline (67%) and ampicillin(50%). CONCLUSION: A. butzleri and A. cryaerophilus cause a small proportion of cases of diarrhoea in humans resident in New Zealand.